The present invention relates to compositions, methods and apparatuses for preserving biological materials. More particularly, the invention relates to compositions, methods and apparatuses for the extended storage of platelets.
Over the last 40 years the need for the therapeutic use of biological materials, such as blood, skin and other tissues, kidneys, hearts, livers and other body organs has increased dramatically. Blood and plasmas components, including red cells, platelets, clotting factors, albumin, and antibodies are isolated and used to treat various bleeding problems. In particular, platelets, essential components of the human blood, are used extensively for assisting in the control of bleeding and replacing functionally defective platelets in patients. For example, platelet transfusions are required by trauma patients who have lost significant amount of blood, patients undergoing chemotherapy that reduces the number of platelets and causes functional defects in remaining platelets, and patients with certain platelet-depleting diseases.
Constituents in whole blood include leukocytes (white blood cells), erythrocytes (red blood cells), thrombocytes, platelets and plasma. Platelets are not entire cells but small detached cell fragments or xe2x80x9cminicellsxe2x80x9d derived from the cortical cytoplasm of large cells called megakaryocytes in the bone marrow. Platelets comprise an outer membrane and cytoplasm from the megakaryocytes which contain granules, dense bodies, dense tubular system and mitochondria. Platelets adhere specifically to the endothelial cell lining of damaged blood vessels, where they trigger and participate in hemostasis, or clotting, and release inflammatory mediators in response to contact with the endothelial cell lining. Important mediators released by platelets include serotonin and coagulation factors. Vascular breaches are repaired by platelets through adhesion and the response to damage is amplified by platelet secretions resulting in platelet aggregation and fibrin formation, i.e. stabilized clot.
It is very important to preserve platelets after their isolation from the body under conditions not only maintaining the biological activity of platelets but also suitable for clinical use. The average survival time for a platelet in the body after it leaves the bone marrow is 8-10 days. The average expected survival time for circulating platelets is 4-5 days, an average of the entire population. The average survival time for platelets after isolation from the body is about 5 days at room temperature.
The current standard and approved method for platelet storage is in a platelet bag at room temperature and limited to five days. The storage time is presumably limited by a decrease in pH due to increased lactate associated with anaerobic metabolic activity. Furthermore, the bag of platelets in plasma must be constantly in motion on a rocker to prevent aggregation. One of the disadvantages associated with preserving platelets under room temperature is the growth of bacteria in the platelet suspension. Platelets in a suspension stored in a refrigerator, albeit with suppressed bacteria growth, tend to activate upon contacting each other and aggregate.
Several approaches such as cryopreservation (freezing) techniques have yielded an increased number of platelets following storage. However, there is a limitation in the functional capacity and persistence of platelets in circulation that are recovered from such preservation conditions by using these methods. Freezing temperatures require the use of cryoprotectors such as DMSO (dimethyl sulfoxide) (Valeri, Feingold, and Marchionni, Blood, vol. 43, No. 1 (January)1974) and THROMBOSO(trademark) to prevent damage to these biological materials. However, these cryoprotectors are cytotoxic, and typically leave a significant portion of the platelets with either reduced or no functional ability. Moreover, cryoprotectors usually require time-consuming preparation, such as rinsing processes, before the materials can be used, and cryoprotector residues often still remain afterwards. Freezing processes can store erythrocytes for more than 30 days, and leukocytes up to 12 hours only.
Other attempts to preserve platelets have included adding platelets activation inhibitors (Bode, Holme, Heaton and Swanson, Vox Sang, 60: 105-112 (1991); U.S. Pat. No. 5,622,867) or gelatin into the preservation medium (U.S. Pat. No. 2,786,014).
A need continues to exist for a storage system that will store biological materials, particularly platelets, for an extended period of time and still maintains their viability and bioactivity.
The present invention relates to compositions, methods and apparatuses for the extended storage of biological material and, in particular, platelets.
According to one embodiment, a platelet composition suitable for direct transfusion into a patient is provided comprising: a preservation medium comprising plasma and a gel-forming material in a concentration relative to the plasma such that the medium is in a sufficiently fluent state at a first temperature to allow platelets to move within the medium and is in a sufficiently gelatinous state at a second, lower temperature to substantially prevent platelets from moving freely within the medium; and platelets.
According to this embodiment, the first temperature is preferably about 37xc2x0 C. and the second temperature is preferably about 5xc2x0 C.
According to another embodiment, a platelet composition suitable for direct transfusion into a patient is provided comprising: a preservation medium comprising plasma and a gel-forming material in a concentration relative to the plasma such that the medium is in a sufficiently fluent state at a first temperature to allow platelets to move within the medium and is in a sufficiently gelatinous state at a second, lower temperature to substantially prevent platelets from moving freely within the medium; and platelets which have been stored within the preservation medium in a gelatinous state for at least 3 days where at least 50% of the platelets are intact and functional after the at least 3 days.
According to this embodiment, the first temperature is preferably about 37xc2x0 C. and the second temperature is preferably about 5xc2x0 C.
Also according to this embodiment, the platelets may be stored within the preservation medium for at least 5 days, more preferably at least 7 days. Also according to this embodiment, the platelets may be stored within the preservation medium for between 3 and 20 days, more preferably between 5 and 20 days. Longer storage of platelets is also possible.
Also according to this embodiment, the platelets may be stored within the preservation medium at a temperature less than 10xc2x0 C. and preferably between xe2x88x9210xc2x0 C. and 10xc2x0 C. In one variation, the platelets are stored at a temperature between 0xc2x0 C. and 10xc2x0 C. at 1 ATM, more preferably at a temperature between 0xc2x0 C. and 5xc2x0 C. at 1 ATM. In another variation, the platelets are stored within the preservation medium at a temperature between xe2x88x9210xc2x0 C. and 0xc2x0 C. at a pressure greater than 10 ATM, more preferably at a temperature between xe2x88x928xc2x0 C. and xe2x88x922xc2x0 C. at a pressure greater than 10 ATM.
According to another embodiment, a platelet composition suitable for direct transfusion into a patient is provided comprising: a preservation medium comprising plasma and a gel-forming material in a concentration relative to the plasma such that the medium is in a sufficiently fluent state at a first temperature to allow platelets to move within the medium and is in a sufficiently gelatinous state at a second, lower temperature to substantially prevent platelets from moving freely within the medium; and platelets which have been stored within the preservation medium in a gelatinous state for at least 1 day at a pressure of at least 10 ATM and a temperature below 0xc2x0 C. where at least 50% of the platelets are intact and functional after the at least 1 day.
According to this embodiment, the first temperature is preferably about 37xc2x0 C. and the second temperature is preferably about 5xc2x0 C.
Also according to this embodiment, the platelets may be stored within the preservation medium at a pressure of at least 30 ATM, more preferably at least 70 ATM, most preferably at least 200 ATM.
According to this embodiment, the platelets may be stored within the preservation medium for at least 3 days, more preferably at least 5 days and most preferably at least 7 days. Also according to this embodiment, the platelets may be stored within the preservation medium for between 2 and 20 days, more preferably between 3 and 20 days. Longer storage of platelets is also possible.
The present invention also relates to a variety of methods for storing platelets for direct transfusion into a patient. In one embodiment, the method comprises:
forming a fluent platelet composition comprising platelets and a preservation medium including plasma and a gel-forming material in a concentration relative to the plasma such that the medium is in a sufficiently fluent state at a first temperature to allow platelets to move within the medium and is in a sufficiently gelatinous state at a second, lower temperature to substantially prevent platelets from moving freely within the medium;
cooling the fluent preservation medium to form a sufficiently gelatinous state to substantially prevent free movement of the platelets within the preservation medium; and
storing the platelets within the preservation medium in a gelatinous state for at least 3 days where at least 50% of the platelets are intact and functional after the at least 3 days.
According to this embodiment, the first temperature is preferably about 37xc2x0 C. and the second temperature is preferably about 5xc2x0 C.
According to this embodiment, the platelets may be stored within the preservation medium for at least 5 days, more preferably at least 7 days. Also according to this embodiment, the platelets may be stored within the preservation medium for between 3 and 20 days, more preferably between 5 and 20 days. Longer storage of platelets is also possible.
Also according to this embodiment, the platelets may be stored within the preservation medium at a temperature less than 10xc2x0 C. and preferably between xe2x88x9210xc2x0 C. and 10xc2x0 C. In one variation, the platelets are stored at a temperature between 0xc2x0 C. and 10xc2x0 C. at 1 ATM, more preferably at a temperature between 0xc2x0 C. and 5xc2x0 C. at 1 ATM. In another variation, the platelets are stored within the preservation medium at a temperature between xe2x88x9210xc2x0 C. and 0xc2x0 C. at a pressure greater than 10 ATM, more preferably at a temperature between xe2x88x928xc2x0 C. and xe2x88x922xc2x0 C. at a pressure greater than 10 ATM.
According to another embodiment, a method is provided for storing platelets for direct transfusion into a patient comprising:
forming a fluent platelet composition comprising platelets and a preservation medium including plasma and a gel-forming material in a concentration relative to the plasma such that the medium is in a sufficiently fluent state at a first temperature to allow platelets to move within the medium and is in a sufficiently gelatinous state at a second, lower temperature to substantially prevent platelets from moving freely within the medium;
cooling the fluent preservation medium to form a sufficiently gelatinous state to substantially prevent free movement of the platelets within the preservation medium; and
storing the platelets within the preservation medium in a gelatinous state for at least 1 day at a temperature below 0xc2x0 C. and at a pressure of at least 10 ATM where at least 50% of the platelets are intact and functional after the at least 1 day.
According to this embodiment, the first temperature is about 37xc2x0 C. and the second temperature is about 5xc2x0 C.
According to this embodiment, the platelets are preferably stored within the preservation medium at a pressure of at least 30 ATM, more preferably at a pressure of at least 70 ATM, most preferably at a pressure of at least 200 ATM.
According to this embodiment, the platelets may be stored within the preservation medium for at least 3 days, more preferably at least 5 days, most preferably at least 7 days. Also according to this embodiment, the platelets may be stored within the preservation medium for between 3 and 20 days, more preferably between 5 and 20 days. Longer storage of platelets is also possible.
In regard to all of the above compositions and methods, it preferred that at least 65% of the platelets are intact and functional after storage, more preferably at least 75% of the platelets, most preferably at least 85% of the platelets.
Also in regard to all of the above compositions and methods, the gel-forming material preferably constitutes between 0.2% and 4% of the preservation medium although the concentration may vary depending on the particular gel-forming material used. Examples of gel-forming material that may be used include, but are not limited to gelatin, agarose, agar, pectin, carob cassia and natural or synthetic water soluble gum such as xanthan gum, konjac gum, guar gum, gum arabic, sodium alginate, carrageenan, irgacanth gum and hydroxyethyl methacrylaic.
Also in regard to all of the above compositions and methods, the preservation medium may further include an energy source. The energy source preferably constitutes between 0 and 5% of the preservation medium, more preferably between 0.25 and 5% of the preservation medium, and most preferably between 0.5 and 5% of the preservation medium. A wide variety of energy sources may be used. Most typically, the energy source is a carbohydrate, such as a sugar. Particular examples of energy sources include glucose, sucrose, mannose, fructose and galactose.
Also in regard to all of the above compositions and methods, the preservation medium may further include water soluble salts. The salt preferably constitutes between 0 and 2% of the preservation medium. Examples of salts include, but are not limited to sodium chloride, potassium chloride, magnesium chloride, sodium phosphate, potassium phosphate and sodium gluconate.
Also in regard to all of the above compositions and methods, the preservation medium may further include an anticoagulant. Examples of anticoagulants that may be used include heparin, citrate dextrose, citrate phosphate dextrose, amantadine, ajoene and ticlopidine.
Also in regard to all of the above compositions and methods, the preservation medium may further include amino acids. Examples of amino acids that may be used include arginine, lysine, aspartate and glutamate.
The present invention also relates to an apparatus for preserving biological materials. In one embodiment, the apparatus includes a chamber having a mouth and a lip, the lip having an inside surface and a top surface, the inside surface and the top surface of the lip meeting at a first radius, the top surface of the lip having a channel. The apparatus also includes a cover configured to mate with and seal the chamber, the cover having a bottom surface, the bottom surface having a protrusion and a sealing structure, the bottom surface of the cover and the protrusion meeting at a second radius, the protrusion being inserted into the mouth of the chamber when the cover is mated with the chamber, the protrusion having a side surface, the side surface of the protrusion and the inside surface of the lip defining a first gap and being substantially parallel when the cover is mated with the chamber, the bottom surface of the cover and the top surface of the lip defining a second gap and being substantially parallel when the cover is mated with the chamber, the second gap having a length greater than a width of the first gap, the sealing structure being inserted into the channel of the lip when the cover is mated with the chamber.